The purpose of this project is to examine gene regulation and expression in the central nervous system, and to optimize gene transfer and antisense technology. The ultimate goal is to achieve neuron~specific expression in cerebellar granule cultures, using highly sensitive reporter gene systems. The luciferase reporter gene system was investigated using electroporation techniques. The luciferase vector, controlled by the Rous sarcoma viral promoter, gave a total of 65,000 light units versus 1,800 for the promoterless vector. Given such high levels of sensitivity, the luciferase vector was used in the construction of neuron~specific vectors. A 386~base pair fragment of the regulatory region of the GAP~43 gene has been identified as controlling neuron~specific expression. This fragment was selected to control the neuron~specific expression of the luciferase gene. Two oligonucleotide primers flanking the 386 bp region were used with rat genomic DNA as template to generate the fragment by the polymerase chain reaction (PCR). Unexpectedly, three fragments were generated, a 386, 500, and 1300 bp fragment. It has been found that these fragments were specific PCR fragments and further characterization of their properties is needed. Construction has begun on an expression vector containing the beta~galactosidase gene in tandem with a multicloning expression cassette. This will allow living transfected cells expressing beta~galactosidase to be identified by fluorescent imaging techniques with simultaneous observation of the cellular effects of the inserted cDNA or antisense construct.